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Purpose

The c-type natriuretic peptide (CNP) signaling pathway is known as a major contributor
to skeletal growth in children. CNP is produced and secreted by both growth plate
and joint chondrocytes, and has a paracrine regulatory effect on cartilage tissue.
CNP increases matrix production by chondrocytes and promotes their proliferation.
In this study, we investigated whether over-expression of CNP in chondrocytes would
be protective of joint cartilage degradation during chronic inflammatory arthritis
in vivo.

Methods

We first developed transgenic mice that over-express CNP (CNPcol2a1TG) in chondrocytes under the control of the collagen 2a1 promoter and enhancer. Then,
we obtained K/BxN TCR transgenic mice from a collaborator and analyzed both transgenic
mice for their joint cartilage clinical and histologic findings over time. We crossed
CNPcol2a1TG mouse with K/BxN TCR transgenic mouse to produce mice with both K/BxN TCR and CNPcol2a1TG backgrounds. The degree of arthritis and cartilage damage in the offspring was analyzed
using a clinical scoring system and two histological scoring systems. Differences
between the scores were analyzed using the Student’s t-test.

Results

Mice that carried the transgene for both CNPcol2a1TG and K/BxN TCR showed less severe clinical and histologic arthritis findings in the
joint cartilage compared to wild type littermates. Between the ages of 6-14 weeks,
the average arthritis score of K/BxN TCR transgenic mice that over-expressed CNP was
4.37 ±1.38 (n=8), while the average arthritis score of K/BxN TCR arthritic mice of
the same age was 8.66 ±3.26 (n=14), (p<0.05). Histological staining and morphometry
did not show any evidence of cartilage degradation in the joint cartilage of CNPcol2a1TG mice. The knee and ankle cartilage of CNPcol2a1TG mice was thick and showed increased proteoglycan content by Safranin-O staining.
However, the double-transgenic offspring mice (K/BxN/CNPcol2a1TG) developed less cartilage damage and less chondrocyte disorganization while still
developing inflammatory changes (pannus) in the synovium, similar to the K/BxN TCR
mice. We adapted the ICRS histological scoring system and gave scores to the knee
joint cartilage of the 8-week-old male mice. K/BxN mice (n=7) scored significantly
lower for both chondrocytic cell distribution (III) and chondrocyte matrix content
(II) (p<0.001 and p<0.05, respectively) than the (K/BxN/ CNPcol2a1TG mice (n=12).

Conclusion

K/BxN TCR arthritic mice over-expressing CNP did not have joint cartilage damage due
to chronic inflammation. We conclude that excess paracrine production of CNP in the
joint cartilage of double-transgenic K/BxN/CNPcol2a1TG arthritic mice was able to overcome the effects of pro-inflammatory cytokines on
joint cartilage in vivo. CNP and the effector molecules of CNP signaling pathway may have therapeutic potential
in protecting cartilage homeostasis during chronic inflammatory arthritis.